ENHANCED STIMULATION OF FOLLICLE MATURATION AND OVULATORY POTENTIAL BY LONG-ACTING FOLLICLE-STIMULATING-HORMONE AGONISTS WITH EXTENDED CARBOXYL-TERMINAL PEPTIDES
ENHANCED STIMULATION OF FOLLICLE MATURATION AND OVULATORY POTENTIAL BY LONG-ACTING FOLLICLE-STIMULATING-HORMONE AGONISTS WITH EXTENDED CARBOXYL-TERMINAL PEPTIDES
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DOI：
10.1210/en.131.6.2514
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发表时间：
1992-12-01
期刊：
影响因子：
4.8
通讯作者：
HSUEH, AJW
中科院分区：
文献类型：
--
作者：
LAPOLT, PS;NISHIMORI, K;HSUEH, AJW
颗粒细胞分化和卵泡成熟的诱导依赖于FSH的刺激作用。我们最近的研究使用重组DNA技术将hCG β亚基的羧基末端肽（CTP）融合到FSH β亚基的羧基末端。所得到的FSH类似物具有与野生型FSH（WT-FSH）相同的体外受体结合和生物活性，但循环半衰期增加。本研究进一步检查了添加一种（FSH-CTP 1）或两种（FSH-CTP 2）CTP的FSH促进颗粒细胞分化和卵泡排卵潜力的能力。WT-FSH、FSH-CTP 1和FSH-CTP 2由用共同α亚基和各自β亚基转染的中国仓鼠卵巢细胞产生。通过RIA定量激素浓度，并通过放射性配体受体测定确认相对水平。FSH-CTP 1和FSH-CTP 2都保留了完整的FSH受体结合活性，但不结合LH受体。为了比较体内生物活性，未成熟雌激素致敏的雌性大鼠在0和24 h接受FSH或激动剂的ip注射。在48 h时，1.0和3.0 IU/天的FSH-CTP 1或FSH-CTP 2诱导卵巢重量的实质性刺激（高达2.5倍），而1.8倍的刺激需要更高剂量（10 IU/天）的WT-FSH。尽管FSH-CTP 1和FSH-CTP 2的体内效力相似，但FSH-CTP在诱导颗粒细胞芳香化酶活性和LH受体方面的效力比WT-FSH高约10倍。我们进一步减少了激素给药的频率。48小时后，单次腹腔注射FSH-CTP 1剂量增加（1-10 IU）导致颗粒细胞芳香酶活性和LH受体含量呈剂量依赖性增加。虽然单次注射（10 IU）WT-FSH没有效果，但间隔12小时注射4次2.5 IU的WT-FSH总剂量相同。为了测试卵泡的排卵潜力，大鼠接受单次注射FSH-CTP 1，52 h后接着注射5 IU hCG以诱导排卵。虽然hCG在接受单次剂量（10 IU）WT-FSH的雌性中未诱导排卵，但在3和10 IU FSH-CTP 1致敏的动物中分别发现20 +/- 2和43 +/- 5个排卵卵子/大鼠。由于每天两次注射WT-FSH（2.5 IU/注射）也增加了卵巢的排卵潜力，因此FSH-CTP 1的有效性增强似乎与其循环半衰期增加有关。本研究结果证实了长效人FSH激动剂与延长的CTP对卵泡成熟和排卵潜能的刺激。这些具有增加的体内效力的重组FSH激动剂的可用性应提供诱导卵泡发育和排卵潜能的改进的手段以及阐明CG β的CTP在体内激素加工中的作用的模型。
The induction of granulosa cell differentiation and follicle maturation is dependent upon the stimulatory actions of FSH. Our recent studies used recombinant DNA technology to fuse the carboxyl-terminal peptide (CTP) of hCG beta-subunit to the carboxyl-terminus of the FSH beta-subunit. The resulting FSH analog has identical in vitro receptor-binding and biological activities as wild-type FSH (WT-FSH), but an increased circulating half-life. The present studies examined further the ability of FSH with one (FSH-CTP1) or two (FSH-CTP2) appended CTPs to promote granulosa cell differentiation and follicle ovulatory potential. WT-FSH, FSH-CTP1, and FSH-CTP2 were produced from Chinese hamster ovary cells transfected with the common alpha-subunit and respective beta-subunit. Hormone concentrations were quantitated by RIA, and relative levels confirmed by radioligand receptor assay. Both FSH-CTP1 and FSH-CTP2 retained full FSH receptor-binding activity, but did not bind LH receptors. To compare in vivo bioactivity, immature estrogen-primed female rats received ip injections of FSH or the agonists at 0 and 24 h. At 48 h, substantial stimulation (up to 2.5-fold) of ovarian weight was induced by 1.0 and 3.0 IU/day FSH-CTP1 or FSH-CTP2, whereas a higher dose (10 IU/day) of WT-FSH was required for an 1.8-fold stimulation. Although the in vivo potencies of FSH-CTP1 and FSH-CTP2 were similar, FSH-CTPs were about 10-fold more potent than WT-FSH in inducing granulosa cell aromatase activity and LH receptors. We further reduced the frequency of hormone administration. Increasing doses (1-10 IU) of a single ip injection of FSH-CTP1 resulted in dose-dependent increases in granulosa cell aromatase activity and LH receptor content 48 h later. Although a single injection (10 IU) of WT-FSH had no effect, the same total dose of WT-FSH administered as four 2.5-IU injections 12 h apart was effective. To test the ovulatory potential of ovarian follicles, rats received a single injection of FSH-CTP1, followed 52 h later by 5 IU hCG to induce ovulation. Although hCG did not induce ovulation in females receiving a single dose (10 IU) of WT-FSH, 20 +/- 2 and 43 +/- 5 ovulated ova/rat were found in animals primed with 3 and 10 IU FSH-CTP1, respectively. Because twice daily injections of WT-FSH (2.5 IU/injection) also increased the ovulatory potential of the ovary, the enhanced effectiveness of FSH-CTP1 appears to be related to its increased circulating half-life. The present results demonstrated the stimulation of follicle maturation and ovulatory potential by long-acting human FSH agonists with extended CTPs. The availability of these recombinant FSH agonists with increased in vivo potency should provide an improved means of inducing follicle development and ovulatory potential as well as a model to elucidate the role of CTP of the CGbeta in hormone processing in vivo.